鎬х棶鑿滆姳

銆€銆€鐢鋒€х殑鐝嶇彔鐘墮槾鑼庝笜鐤圭棶鍜屽コ鎬х殑鍋囨€ф箍鐤ｄ笌灝栭攼婀跨枺鏈変綍涓嶅悓錛?銆€銆€鐢鋒€х殑鐝嶇彔闃磋寧涓樼柟鐥呭拰濂蟲€х殑鍋囨箍鐤ｄ笌灝栭攼婀跨枺涓昏鍖哄埆濡備笅錛?銆€銆€錛?錛夌柧鐥呯殑鑼冨洿涓嶅悓 銆€銆€鐝嶇彔鐘墮槾鑼庝笜鐤圭棶鍜屽コ鎬у亣鎬ф箍鐤ｆ槸涓€縐嶈壇鎬х毊鑲ゅ彂鑲蹭笉鍏ㄧ殑鐘舵€侊紝鍦ㄦ甯鎬漢緹や腑鏈?0%鐨勪漢鏈夋鐥呫€傛嵁璋冩煡錛屽湪澶勫コ鍜屾病鏈夎繃鎬ф帴瑙︾殑鐢鋒€т腑錛屾湰鐥呯殑鎮ｇ棶鐜囦篃寰堥珮銆傛湰鐥呬笉鏄敱鎬ф帴瑙﹁€屾劅鏌撶殑錛屼笉灞炰簬鎬х棶鑼冨洿銆傝€屽皷閿愭箍鐤ｏ紝鏄€氳繃鎬ф帴瑙﹁€屾劅鏌撶殑錛屾槸鎬х棶涓殑涓€縐嶃€?銆€銆€錛?錛夎〃鐜頒笉鍚?銆€銆€鐢鋒€х弽鐝犵姸闃磋寧涓樼柟鐥呯殑鐗圭偣錛屾槸鍦ㄦ部鍐犵姸娌熷鍒嗗竷鐨勫皬綾崇矑澶у皬鐨勪笜鐤廣€傚コ鎬у亣鎬ф箍鐤ｆ槸鍦ㄥ皬闃村攪鍐呬晶鍛堝縐版€х殑銆佹潯緔㈠艦鎸囩姸涓樼柟錛屽垎甯冪瀵嗭紝涔熷彲鍛堥奔瀛愮姸銆傛湰鐥呭彲鏃犱換浣曚笉閫傛劅錛屼笉闇€娌葷枟錛屾棤浼犳煋鎬с€傝€屽皷閿愭箍鐤ｆ槸鍦ㄧ敺濂崇殑澶栫敓孌栧櫒澶勬暎鍦ㄧ殑錛屽憟鑿滆姳鐘剁毊鑲ょ枺鐘剁墿錛屽垎甯冩棤瑙勫緥鎬э紝鍙湁鎼旂棐鎰燂紝鏈夊紓鍛熾€傞槾閬撳唴鐢熼暱鐨勫皷閿愭箍鐤ｅ彲鑷鍑鴻錛屾湁鑷韓浼犳煋鎬э紝鏈粡娌葷枟鏈棶鍙戶緇彂灞曪紝 鍏舵暟鐩秺鏉ヨ秺澶氾紝鐤ｄ綋閫愭笎澧炲ぇ銆?銆€銆€錛?錛夋不鐤楃洰鐨勪笉鍚?銆€銆€鐢鋒€х弽鐝犵姸闃磋寧涓樼柟鐥呭拰濂蟲€у亣鎬ф箍鐤ｅ彲鏃犱換浣曚笉閫傛劅錛屼笉闇€娌葷枟錛涜€屽皷閿愭箍鐤ｆ槸鎬т紶鎾柧鐥咃紝浼犳煋鎬у己錛屽繀欏繪不鐤楋紝鍚﹀墶鍚庢灉鏇村姞涓ラ噸銆?銆€銆€涓村簥涓婂鏄撳皢鐢鋒€х弽鐝犵姸闃磋寧涓樼柟鐥呭拰濂蟲€у亣鎬ф箍鐤ｈ璿婁負灝栭攼婀跨枺錛岃€屾墸涓婃€х棶鐨勫附瀛愶紝鍥犳錛屽尰鐢熶竴瀹氳璿婃柇娓呮銆傜棶浜鴻縐瀬閰嶅悎錛岃鐪熷洖絳斿尰鐢熺殑闂瘖錛岃鍋氬埌鏃㈣鎺掗槦鎬х棶鐨勫彲鑳斤紝鍚屾椂涔熶笉瑕佽鎬х棶婕忚瘖銆?銆€銆€ 銆€銆€娌葷枟灝栭攼婀跨枺甯哥敤鑽墿 銆€銆€鍏鋒湁瀵圭枺浣撶牬鍧忔€х殑鑽墿閮借兘鐢ㄤ簬灝栭攼婀跨枺鐨勬不鐤椼€備笅闈粙緇嶅嚑縐嶅父鐢ㄧ殑娌葷枟灝栭攼婀跨枺鐨勮嵂鐗╋紝渚涜鑰呭弬鑰冦€?銆€銆€(1)5錛?-Fu杞啅銆€5-Fu鏄竴縐嶅父鐢ㄧ殑鎶楄偪鐦よ嵂鐗╋紝瀹冭兘鎶戝埗緇嗚優鐨勫孌栵紝鍔溳噺杈冧負娓╁拰錛屼竴鑸渶娌葷枟2鍀?鍛ㄣ€?銆€銆€(2)30錛呬笁姘唻閰搞€€鏄竴縐嶅己閰革紝鍙敤鏉ョ儳鐏肩枺浣擄紝姣忓懆鐢ㄨ嵂2嬈★紝瑕佹敞鎰忎繚鎶ゅ懆鍥寸毊鑲ゅ拰綺樿啘銆?銆€銆€(3)浜夊厜闇夌礌銆佸鉤闃抽湁緔犲厬鐢熺悊鐩愭按錛屽娑傛偅澶勶紝姣忓懆鐢ㄨ嵂2鍀?嬈°€?銆€銆€(4)3錛呯殑鑲戒竵鑳洪湝銆€澶栨惤鏈嵂錛屾瘡鏃?嬈★紝鏃犱笉鑹壇浣滅敤錛屽皻鍙敤浜庨槾閬撳唴鐨勫皷閿愭箍鐤ｃ€傚姏閲忔俯鍜岋紝闇€闀挎湡娌葷枟銆?銆€銆€(5)鐤ｈ劚嬈ｏ紙楝艱嚰姣掔礌婧舵恫錛夈€€涓烘柊鐨勯珮鏁堢殑娌葷枟灝栭攼婀跨枺鐨勮嵂鐗╋紝鏃犲埡嬋€鎬э紝澶栫敤灞€閮紝姣忔棩1嬈★紝榪炵畫3鍀?澶╁嵆鍙€?銆€銆€(6)20錛呯殑瓚沖彾鑽夐叝閰娿€€姣忓懆鐢?鍀?嬈°€傛湰鑽湁鍓綔鐢紝瑕佹敞鎰忎繚鎶ゅ懆鍥寸毊鑲ょ矘鑶滐紝娑傝嵂2鍀?灝忔椂鍚庢礂鍘匯€傛湁鑷寸暩浣滅敤錛屽瓡濡囩鐢ㄣ€?銆€銆€灝栭攼婀跨枺鐨勫父鐢ㄦ不鐤楁柟娉?銆€銆€鐩墠娌葷枟灝栭攼婀跨枺涓昏鏄互澶栨不涓轟富錛屽唴娌諱負杈呫€傚娌葷殑鏂規硶寰堝錛屾湁鑽墿娌葷枟銆佸喎鍐繪不鐤椼€佹縺鍏夋不鐤椼€佸井娉㈡不鐤椼€佺數鐑ф不鐤椼€佹墜鏈不鐤楃瓑銆傚畠浠悇鏈夊叾閫傚簲鐥囷紝鍏舵不鎰堢巼鍦?0%鍀?0錛呬箣闂淬€傝嵂鐗╂不鐤楁柟渚匡紝鏄撲簬鎿嶄綔錛屼絾涓€鑸渶瑕侀噸澶嶆暟嬈★紝鑰屼笖鏈変簺鑽墿鏈夌伡浼ょ矘鑶滅殑鍙兘銆傛縺鍏夋不鐤椼€佸喎鍐繪不鐤椼€佸井娉㈡不鐤楅渶瑕佹湳鑰呮湁楂樿秴鐨勬妧鑹猴紝鍑嗙‘鍦扮牬鍧忕棶鍙樼粍緇囥€?銆€銆€錛戙€佸喎鍐葷枟娉?鏄竴縐嶇毊鑲ょ甯哥敤鐨勬不鐤楁柟娉曪紝瀹冩槸浠ユ恫姘垨浜屾哀鍖栫⒊騫插啺鍐峰喕鐨偆鐥呮崯錛屼嬌鐨偆鎹熷灞€閮ㄦ按鑲褲€佸潖姝夥紝浠ヨ揪鍒版不鐤楃殑鐩殑銆傚皷閿愭箍鐤ｆ槸鐢變簬灝栭攼婀跨枺鐥呮瘨鐨勬劅鏌擄紝瀵艱嚧鐨偆綺樿啘鐨勮壇鎬у鐢熴€傚畠鏈夊ぇ閲忕殑灝忚綆★紝澧炴畺榪呴€熴€傜敤鍐峰喕鐨勬柟娉曞彲浣垮皷閿愭箍鐤ｅ唴緇撳啺錛屽艦鎴愮粍緇囧矓閮ㄧ殑楂樺害姘磋偪錛屼粠鑰岀牬鍧忕枺浣撱€傚喎鍐繪不鐤楃殑鏈€澶уソ澶勬槸灞€閮ㄤ笉鐣欑棔榪癸紝娌繪剤鐜囩害70錛呭乏鍙熾€傚彲鐢ㄥ柗闆炬硶鎴栫洿鎺ユ帴瑙︽硶錛岄€傜敤浜庡鉤鍧﹁緝灝忕殑鐤ｄ綋銆備竴鑸瘡鍛ㄥ喎鍐?嬈★紝榪炵畫2鍀?嬈°€傘€€ 銆€銆€錛掋€佹縺鍏夋不鐤?鏄毊鑲ゆ€х棶縐戝父鐢ㄧ殑娌葷枟灝栭攼婀跨枺鐨勬柟娉曘€傚畠鐨勭壒鐐規槸瑙佹晥蹇紝鍦ㄦ不鐤楃殑褰撴椂錛岀枺浣撳嵆鍙劚钀姐€傚父鐢ㄧ殑鏄簩姘у寲紕蟲縺鍏夛紝鐑х伡鐤ｄ綋錛屼竴鑸?嬈″嵆鍙嬌鐤ｄ綋鑴辮惤銆備絾鏄 紝鐢變簬嬋€鍏夌殑鍏夋潫榪囦簬闆嗕腑鐨勫師鏁咃紝嬋€鍏夋不鐤楀彧閫傜敤浜庣枺浣撹緝灝忕殑灝栭攼婀跨枺錛屽鐤ｄ綋杈冨ぇ錛屾縺鍏夋不鐤楀緢瀹規槗澶嶅彂銆?銆€銆€錛撱€佸井娉㈡不鐤?鏄繘鍏?0騫翠唬鍚庡叴璧風殑娌葷枟鏂規硶銆傚畠鐨勫師鐞嗘槸鍒╃敤寰嘗鐨勯珮棰戦渿鍔紝浣跨枺浣撳唴閮ㄦ按鍒嗚捀鍙戯紝鍧忔鑴辮惤銆傚井娉㈡不鐤楃殑鐗圭偣鏄紝鐤ｄ綋鐮村潖褰誨簳錛屼笉鏄撳鍙戯紝浣嗗壋闈㈡仮澶嶈緝鎱紝瀹規槗緇у彂鎰熸煋銆傛墍浠ュ井娉㈡不鐤楃壒鍒€傜敤浜庢不鐤楃枺浣撹緝澶х殑錛屽絝嬨€佹暎鍦ㄧ殑錛屽悓鏃惰榪滅鍖呯毊緋誨甫鐨勫皷閿愭箍鐤ｏ紝浠ラ槻鎹熶激鍖呯毊緋誨甫錛屽獎鍝嶇敺鎬х殑鎬у姛鑳姐€?銆€銆€錛斻€佺數鐑х伡 鏄緝鑰佺殑涓€縐嶆不鐤楁柟娉曘€傝繃鍘葷毊鑲ょ鍖誨笀甯哥敤楂橀鐢靛垁鎴栫數鐑欓搧鐑х伡瀵誨父鐤ｃ€佽壊緔犵棧鍙婂叾浠栫毊鑲よ禈鐢熺墿銆傚畠鐨勭壒鐐規槸鎿嶄綔綆€鍗曪紝瑙佹晥蹇€傞珮棰戠數鍒€鑳界洿鎺ュ垏闄ゅ拰騫茬嚗鐤ｄ綋錛屾不鐤椾篃杈冨交搴曪紝浣嗘槸錛岀數鐑х伡鐤潰鎰堝悎杈冪紦鎱€傜患涓婃墍榪幫紝鐢電儳鐏兼不鐤楋紝鍙敤浜庝換浣曞皷閿愭箍鐤ｇ殑娌葷枟錛屼絾鏄鏂芥湳鑰呯殑鎶€鏈姹傝緝楂橈紝鐑х伡澶繃鎴栦笉瓚抽兘鏄湁瀹崇殑銆傚悓鏃惰娉ㄦ剰鏃犺弻鎿嶄綔錛岄闃叉劅鏌撱€?銆€銆€錛曘€佹墜鏈不鐤?灝栭攼婀跨枺涓€鑸笉涓誨紶鎵嬫湳鍒囬櫎錛屽洜涓烘墜鏈不鐤楀悗錛屽皷閿愭箍鐤ｅ緢瀹規槗澶嶅彂錛屼嬌娌葷枟澶辮觸銆備絾瀵瑰甫钂傜殑杈冨ぇ鐨勭枺浣撳彲鑰冭檻鎵嬫湳鍒囬櫎銆傛湁鐨勬偅鑰呭皷閿愭箍鐤ｇ敓闀胯繃浜庤繀閫燂紝鎴栧ぇ濡傝彍鑺憋紝鍏朵粬娌葷枟鍗佸垎鍥伴毦錛屽彲鑰冭檻鎵嬫湳娌葷枟銆備負闃叉澶嶅彂錛屾湳鍚庨厤鍚堝叾浠栨不鐤椼€?

瀹為獙瀹ゆ鏌?銆€銆€涓€銆侀唻閰哥櫧璿曢獙 銆€銆€鐢?-5%閱嬮吀澶栨秱鐤ｄ綋2-5鍒嗛挓錛岀棶鐏墮儴浣嶅彉鐧界◢闅嗚搗錛岃倹闂ㄧ棶鎹熷彲鑳介渶瑕?5鍒嗛挓銆傛湰璿曢獙鐨勫師鐞嗘槸錏嬬櫧璐ㄤ笌閰稿嚌鍥哄彉鐧界殑緇撴灉錛孒PV鎰熸煋緇嗚優浜х敓鐨勮錏嬬櫧涓庢甯哥殑鏈劅鏌撲笂鐨粏鑳炰駭鐢熺殑涓嶅悓錛屽彧鏈夊墠鑰呮墠鑳借閱嬮吀鑴辮壊銆傞唻閰哥櫧璿曢獙媯€嫻婬PV鐨勬晱鎰熸€у緢楂橈紝瀹冩瘮甯歌媯€嫻嬭瀵熺粍緇囧鍙樺寲榪樺ソ銆備絾鍋跺皵鍦ㄤ笂鐨鍘氭垨澶栦激鎿︾牬鐥呬緥涓嚭鐜板亣闃蟲€с€佸亣闃蟲€у彉鐧借抗璞℃樉寰楃晫闄愪笉娓呭拰涓嶈鍒欍€傜編鍥紺DC鎻愮ず錛岄唻閰哥櫧璿曢獙騫朵笉鏄壒寮傝瘯楠岋紝涓斿亣闃蟲€ц緝甯歌銆?銆€銆€浜屻€佸厤鐤粍緇囧媯€鏌?銆€銆€甯哥敤榪囨哀鍖栫墿閰舵姉榪囨哀鍖栫墿閰舵柟娉曪紙鍗砅AP錛夛紝鏄劇ず婀跨枺鍐呯殑鐥呮瘨錏嬬櫧錛屼互璿佹槑鐤ｆ崯瀹充腑鏈夌棶姣掓姉鍘熴€侶PV錏嬬櫧闃蟲€ф椂錛屽皷閿愭箍鐤ｇ殑嫻呰〃涓婄毊緇嗚優鍐呭彲鍑虹幇娣＄孩鑹茬殑寮遍槼鎬у弽搴斻€?銆€銆€涓夈€佺粍緇囧寲瀛︽鏌?銆€銆€鍙栧皯閲忕棶鎹熺粍緇囧埗鎴愭秱鐗囷紝鐢ㄧ壒寮傛姉浜虹被涔沖ご鐦ょ棶姣掔殑鎶椾綋浣滄煋鑹層€傚鐥呮崯涓湁鐥呮瘨鎶楀師錛屽墶鎶楀師鎶椾綋緇撳悎銆傚湪榪囨哀鍖栫墿閰舵姉榪囨哀鍖栫墿閰訛紙PAP錛夋柟娉曚腑錛屾牳鍙鏌撴垚綰㈣壊銆傛娉曠壒寮傛€у己涓旇緝榪呴€燂紝瀵矽瘖鏂湁甯姪銆?銆€銆€鍥涖€佺棶鐞嗘鏌?銆€銆€涓昏涓鴻鍖栦笉鍏紝媯樺眰楂樺害鑲ュ帤錛屼鉤澶寸槫鏍峰鐢燂紝琛ㄧ毊紿佸鍘氾紝寤墮暱錛屽叾澧炵敓紼嬪害鍙技鍋囨€т笂鐨槫鏍楓€傚埡緇嗚優鍜屽熀搴曠粏鑳炲苟鏈夌溮褰撴暟閲忕殑鏍稿垎瑁傘€侀浼肩檶鍙樸€備絾緇嗚優鎺掑垪瑙勫墶錛屼笖澧炵敓涓婄毊鍜岀湡鐨箣闂寸晫闄愭竻妤氥€傚叾鐗圭偣涓虹矑灞傚拰鍒哄眰涓婇儴緇嗚優鏈夋槑鏄劇殑絀烘場褰㈡垚銆傛縐嶇┖娉＄粏鑳炶緝姝ｅ父澶э紝鑳炴祮鐫€鑹叉貳銆佷腑澶湁澶ц€屽渾錛屾繁鍡滅⒈鎬х殑鏍搞€傞€氬父鐪熺毊姘磋偪銆佹瘺緇嗚綆℃墿寮犱互鍙婂懆鍥磋緝鑷村瘑鐨勬參鎬х値鎬ф蹈娑︺€侭ushke-loewenstein宸ㄥぇ鍨嬪皷閿愭箍鐤ｏ紝琛ㄧ毊鏋佸害鍚戜笅鐢熼暱錛屼唬鏇誇簡鍏朵笅闈㈢殑緇勭粐銆佹槗涓庨碁鐘剁粏鑳炵溮娣鳳紝鏁呴』澶氭媧繪銆傝嫢鏈夌紦鎱㈠彂灞曚箣鍊懼悜錛?鍒欎負涓€縐嶄綆搴︽伓鍙樼殑榪囩▼錛屽嵆鎵€璋撶枺鐘剁檶銆?銆€銆€浜斻€佸熀鍥犺瘖鏂?銆€銆€榪勪粖錛孒PV闅句簬鐢ㄤ紶緇熺殑鐥呮瘨鍩瑰吇鍙婅娓呭鎶€鏈嫻嬶紝涓昏瀹為獙璿婃柇鎶€鏈槸鏍擱吀鏉備氦銆傝繎騫存潵鍙戝睍鐨凱CR鏂規硶鍏鋒湁鐗瑰紓銆佹晱鎰熴€佺畝渚褲€佸揩閫熺瓑浼樼偣錛屼負HPV媯€嫻嬪紑杈熶簡鏂伴€斿緞銆?銆€銆€錛堜竴錛夋爣鏈殑閲囬泦鍙婂鐞?銆€銆€1錛庢爣鏈殑閲囬泦鍜岄澶勭悊錛氱敤鍒澘鎴栫敓鐞嗙洂姘存蹈娑︾殑媯夋浠庨槾閬撳拰瀹澶栧彛鍙栧垎娉岀墿鍜岀粏鑳炪€傚湪浣滅粏鑳炲媯€鏌ョ殑鍚屾椂錛屽皢鏍囨湰鏀懼靉5ml鍚湁0.05%紜煶姹炵殑PBS涓紝鐢≒BS紱誨績錛?000g錛?0min錛夋礂娑?嬈★紝娌夌Н緇嗚優閲嶆偓浜?mlPBS涓紝鍙?.5ml緇嗚優鎮恫鎶芥彁DNA銆?銆€銆€2錛庢爣鏈牳閰哥殑鎻愬彇錛氭寜1浣撶Н緇嗚優鎮恫鍔?0鍊嶄綋縐殑緇嗚優瑁傝В娑詫紙10mmol/L Tris-HCl錛宲H 7.4錛?0mmol/L EDTA錛?50mmol/L NaCl錛?.4%SDS錛?.0mg/ml錏嬬櫧閰禟錛?7鈩冧笅澶勭悊榪囧;涓旂瓑浣撶Н閰?姘豢(1:1)錛屾隘浠?寮傛垔閱囷紙24:1錛夊悇鎶芥彁2嬈★紱鍔?/10浣撶Н3mol/L NaAc錛坧H 5.2錛夊強2.5鍊嶄綋縐棤姘翠箼閱囩疆-20鈩?2h鎴栬繃澶滄矇娣€DNA錛涘姞1浣撶Н涔欓唶媧楁釘1嬈★紱鐢?0渭l鍚玆NA閰?100渭g/ml)鐨凾E娑?10mmol/L Tris-HCl錛宲H 8.0錛?.0mmol/L EDTA)婧惰ВDNA錛?7鈩冩俯鑲?0min銆?銆€銆€錛堜簩錛塒CR鎵╁ 銆€銆€1錛?寮曠墿璁捐鍜屽悎鎴愶細HPV鍩哄洜緇勫彲鍒嗕負鏃╂湡鍖猴紙E錛夊拰鏅氭湡鍖猴紙L錛夛紝姣忓尯鍚竴緋誨垪寮€鏀捐鐮佹鏋訛紙ORF錛夈€傚簭鍒楀垎鏋愯〃鏄庯紝鍚勫瀷HPV鐨勯潪緙栫爜鍖哄強E1銆丒6銆丒7鍜孡1鍖哄潎鏈変繚瀹堝簭鍒椼€侻anos絳変粠HPVL1鍖轟腑閫夋嫨淇濆畧搴忓垪璁捐鍚堟垚寮曠墿MY11鍜孧Y09瑙佽〃1錛岃寮曠墿涓嶩PV 6銆?1銆?6銆?8鍙?3鍨嬫湁浜掕ˉ搴忓垪錛屼篃鍙墿澧炲叾瀹冨瀷HPV銆?銆€銆€Manos絳夎璁＄殑HPVL1閫氱敤寮曠墿 銆€銆€MY11錛欸CMCAGGGWCATAAYAATGG 銆€銆€MY09錛欳GTCCMARRGGAWACTGATC 銆€銆€琛? HPV鍨?MY11鐨勭1涓》鍩轟綅緗?MY09鐨勭1涓》鍩轟綅緗?PCR浜х墿闀垮害錛坆p錛?6 6722 7170 448 11 6707 7155 448 16 6584 7035 451 18 6558 7012 454 33 6539 6987 448 M=A+C錛?R=A+G錛學=A+T錛孻= C+T 銆€銆€琛?鍒楄堪浜哅anos絳夎璁＄殑瀵℃牳鑻烽吀鎺㈤拡銆傚叕鐢ㄦ販鍚堟帰閽堝簭鍒楅€夎嚜HPVL1鍖轟腑鍙︿竴淇濆畧搴忓垪錛屽彲涓?MY11鍜孧Y09寮曠墿浜х敓鐨勫縐岺PVL1 PCR鎵╁浜х墿鏉備氦錛氬瀷鐗瑰紓鎺㈤拡鍙笌鐩稿簲HPV鍨嬫湁浜掕ˉ搴忓垪錛岀敤浜庢鍑虹殑HPV鍒嗗瀷銆?銆€銆€琛? Manos絳夎璁＄殑HPVL1 PCR浜х墿鎺㈤拡 鍏敤娣峰悎鎺㈤拡 GP1 CTGTTGTTGATACTACACGCAGTAC GP2 CTGTGGTAGATACCACWCGCAGTAC 絎竴涓》鍩轟綅緗?HPV6 6771 HPV11 6757 HPV16 6631 HPV18 6607 HPV33 6588 鍨嬬壒寮傛帰閽?br /> 鎺㈤拡 HPV鍨?搴忓垪 鍩哄洜浣嶇疆 MY12 HPV6 CATCCGTAACTACATCTTCCA 6813鈥?833 MY13 HPV11 TCTGTGTCTAAATCTGCTACA 6800鈥?820 MY14 HPV16 CATACACCTCCAGCACCTAA 6926鈥?945 MY74 HPV18 GGATGCTGCACCGGTCTGA 6905鈥?922 MY16 HPV33 CACACAAGTAACTAGCTACAG 6628鈥?648 銆€銆€ H=A+C+T錛孯=A+G錛學=A+T錛孻=C+T 銆€銆€2錛嶱CR鍙嶅簲璿曞墏錛歍aq DNA鑱氬悎閰訛紙2U/ml錛夈€?0mmol/L dNTP璐娑詫紙dATP銆乨CTP銆乨GTP銆乨TTP鍚?0mmol/L錛夛紝10脳PCR緙撳啿娑?500mmol KCl銆?0mmol/L MgCl2銆?00mmol/L Tris-HCl銆乸H 8.5)錛?00渭mol/L MY11鍜孧Y09璐娑詫紝鐏弻鐨勭幓鐠冭捀棣忓櫒鍒跺鐨勮捀棣忔按銆?銆€銆€3錛嶱CR鎵╁鏂規硶鍜岀▼搴忥細浠?00渭l PCR鍙嶅簲娑?鐢ㄦ棤鑿?.5ml紜呭寲濉戞枡紱誨績綆′負鍙嶅簲綆¤繘琛屾墿澧炲弽搴斻€?銆€銆€錛?錛夊疄楠屽墠閰嶅埗棰勬販鍙嶅簲璿曞墏騫跺垎瑁呫€傞娣峰弽搴旇瘯鍓傚寘鎷櫎鏍囨湰DNA澶栫殑鍏跺畠鍚勭PCR鍙嶅簲璿曞墏銆傛瘡鏀弽搴旂涓惈鏈夌殑PCR鍙嶅簲璿曞墏瑙佽〃3銆?銆€銆€琛? 姣忔敮鍙嶅簲綆′腑鐨凱CR鍙嶅簲璿曞墏 銆€銆€鍙嶅簲璿曞墏 浣撶Н錛埼糽錛?緇堟祿搴?10脳PCR緙撳啿娑?10 1脳PCR緙撳啿娑?dNTP璐娑?2 200渭mol/L姣忕dNTP MY11璐娑?0.5 500渭mol/L MY09璐娑?0.5 500渭mol/L Taq DNA鑱氬悎閰?0.5 2.5渭 鐏弻钂擱姘?75.5 鎬昏 90 銆€銆€錛?錛夊悇鍙嶅簲綆′緷嬈″姞鍏?0渭l鏍囨湰鍜?0渭l棰勬販鍙嶅簲璿曞墏銆?銆€銆€錛?錛夊姞鍏?0-100渭l鐭寵湣娌癸紝鍦ㄥ彴寮忕蹇冩満涓婂揩閫熺蹇冩暟縐掗挓錛屼嬌鍚勫弽搴旇瘯鍓傛敹闆嗕簬娌瑰眰涓嬨€傜洰鍓嶏紝PCR璿曞墏宸插晢鍝佸寲錛屽弽搴斾綋縐負25渭l銆備嬌鐢ㄦ椂鍙姞鍏ユ爣鏈珼NA鍗沖彲銆?銆€銆€錛?錛夊皢鍙嶅簲綆＄疆PCR鎵╁浠笂錛屽驚鐜弬鏁頒負95鈩?30s錛?5鈩?40s錛?2鈩?50s寰幆35嬈★紝鏈€鍚?2鈩冨歡浼?min銆?銆€銆€4錛庢瘡嬈¤瘯楠屽簲璁鵑槼鎬у強闃存€у鐓с€備互杞芥湁HPV鐨勯噸緇勮川綺掞紙姣忓弽搴斾負100pg錛夋垨鍚湁HPV鐨勭粏鑳炵郴錛堝Caski銆丠eLa錛塂NA涓洪槼鎬у鐓э紝浠ユ棤HPV鐨勪漢緇嗚優緋籇NA涓洪槾鎬у鐓с€?銆€銆€錛堜笁錛夋墿澧炰駭鐗╃殑媯€嫻嬪拰鍒嗘瀽 銆€銆€1錛?鍑濊兌鐢墊吵錛氭墿澧炲弽搴旂粨鏉熷悗錛屽彇鍑哄弽搴旂錛屽喎鍗磋嚦瀹ゆ俯錛屽彇10渭l鎵╁浜х墿鐢?%-7%鑱氫笝鐑叞鑳哄嚌鑳舵垨1.5%鐞艱剛緋栫數娉籌紝婧村寲涔欓敪鏌撹壊錛岀傳澶栧垎鏋愪華涓嬪垎鏋愮粨鏋滐紝鍒嗗瓙閲忕害涓?50bp澶勫嚭鐜版槑鏄劇殑DNA甯︺€?銆€銆€2錛?鏍擱吀鏉備氦錛氬鏋滃嚌鑳剁數娉蟲棤娓呮鐨凞NA鎴栭渶紜畾DNA甯︾殑鐗瑰紓鎬ф椂錛屽彲鐢ㄦ爣璁扮殑鍏敤娣峰悎鎺㈤拡鍜岋紙鎴栵級鍨嬬壒寮傛帰閽堜綔Southern鍚稿嵃鏉備氦銆佹枒鐐規潅浜ら獙璿併€?銆€銆€鎸夌収鏍囧噯鏂規硶鍒?2P ATP鏍囪鐨勫鏍歌嫹閰告帰閽堬紝闇€杈懼埌綰?07cpm/pmol鐗瑰紓媧繪€с€傛潅浜ゆ恫涓渶鍚湁2脳106-5脳106cpm鎺㈤拡/ml銆傚湪55鈩冪紦鎱㈡尟鑽′笅鏉備氦2-3h錛岄殢鍚庝簬30-55鈩冧笅鐢ㄦ礂娑ゆ恫錛?脳SSC銆?.1%SDS錛夎繀閫熷啿媧楁潅浜よ啘錛岄櫎鍘誨浣欐帰閽堛€傜劧鍚庤繘琛屾礂鑶滐紝鍏舵潯浠朵緷鎵€鐢ㄦ帰閽堣€屽紓錛氬叕鐢ㄦ販鍚堟帰閽堬紝55鈩冩礂鑶?0min錛汳Y12銆丮Y13鍙奙Y16鎺㈤拡錛?6-57鈩冧笅10 min錛屽苟鎹㈡恫閲嶆礂1嬈★紱MY14鍙奧D74鎺㈤拡錛?8-59鈩冧笅10min錛屼害鎹㈡恫閲嶆礂1嬈°€?銆€銆€鐢≒CR鏂規硶媯€嫻婬PV姣旀牳閰告潅浜ゆ柟娉曚紭瓚娿€傚叾鏁忔劅鎬ч珮錛孏P-PCR鏂規硶浠ュ嚌鑳剁數娉崇洿鎺ュ垎鏋愮粨鏋滐紝鍙鍑烘爣鏈腑200涓嫹璐濈殑HPV DNA錛岃嫢浠ユ牳閰告潅浜ゆ嫻婸CR浜х墿錛屾晱鎰熸€ф彁楂橈紝鑳芥鍑?0涓嫹璐濈殑HPV DNA銆?銆€銆€閴翠簬PCR鎶€鏈殑楂樺害鏁忔劅鎬э紝浠ョ敓孌栭亾鑴辮惤緇嗚優涓烘鏉愯凍浠ユ弧瓚寵瘯楠岃姹傦紝閬垮厤浜嗘椿媯€鍙栨潗銆佺爺紓ㄧ粍緇囩箒鏉傛搷浣溿€備竴鑸儏鍐典笅錛孭CR鎵╁浜х墿緇忓嚌鑳剁數娉籌紝瑙傚療浜х敓鐨凞NA鍙洿鎺ヤ綔鍑鴻瘖鏂€傚洜姝わ紝PCR鎶€鏈嫻婬PV瀹為獙鍛ㄦ湡鐭€佺畝渚垮揩閫熴€?

銆€銆€ 銆€銆€鍏嶇柅瀛?銆€銆€瀹誇富瀵笻PV鎰熸煋寮曡搗鐨勫厤鐤簲絳旓紝鍖呮嫭緇嗚優鍏嶇柅涓庝綋娑插厤鐤袱涓柟闈€?銆€銆€涓€銆佺粏鑳炲厤鐤?銆€銆€浜轟綋緇嗚優鍏嶇柅鐘舵€佹槸褰卞搷CA鍙戠敓銆佽漿褰掔殑閲嶈鍩虹涔嬩竴銆傜粏鑳炲厤鐤瘮浣撴恫鍏嶇柅鏇翠負閲嶈銆備複搴婁笂浼寸粏鑳炲厤鐤己闄風殑灝栭攼婀跨枺鎮ｈ€呯毊鐤瑰父鎸佺畫涓嶉€€錛屽叾澶栧懆琛€涓姂鍒舵€緇嗚優鏁板澶歂K緇嗚優鍔熻兘浣庝笅錛宺-騫叉壈緔犲拰鐧界粏鑳炰粙緔?浜х敓鍑忓皯錛岃€屾秷閫€鐤ｇ毊鎹熶腑甯稿父鍑虹幇媧誨寲T緇嗚優鍜孨K緇嗚優鐨勬蹈娑︼紝閮ㄥ垎瑙掓湂緇嗚優HLA-DR闃蟲€с€?銆€銆€鍏嶇柅鎶戝埗鎴栧厤鐤己闄鋒椂錛岀敓孌栧櫒HPV鎰熸煋鍜孒PV鐩稿叧鐤劇棶鐨勫彂鐢熺巼鍧囧鍔犮€傚皷閿愭箍鐤ｄ腑杈呭姪鎬緇嗚優鑰楃錛孋D4/CD8姣斿€煎€掔疆錛屽叾鍊鹼紲1銆傚湪CA鐥呬漢鐨勫鍛ㄨ涓紝鍙戠幇鎶戝埗/緇嗚優姣掓€緇嗚優鐧懼垎鐜囨樉钁楀楂橈紝杈呭姪/璿卞鎬緇嗚優鐨勬瘮鍊煎拰杈呭姪/鎶戝埗T緇嗚優姣斿€煎潎闄嶄綆銆傚棰圕A鍜屽棰堜笂鐨唴鐦ゆ牱鐥呭彉錛圕IN錛夌棶鎹熶腑鏈楁牸緗曠粏鑳炴槑鏄懼噺灝戙€侰A涓璑K緇嗚優 浜х敓r-騫叉壈緔犲拰鐧戒粙緔?2鍑忓皯銆傞矋娓╂牱涓樼柟鐥呭拰鑲溳棬鐢熸畺鍣ㄧ檶鑰呬腑NK緇嗚優瀵瑰惈鏈塇PV-16鐨勮鏈婄粏鑳炵殑婧惰В媧繪€т笅闄嶏紝鍙兘鏄鐤劇棶鐗瑰紓鎬ч澏緇嗚優鐨勮瘑鍒己闄鋒墍鑷達紝瀹CA鐨勮鏈婄粏鑳炰笉琛ㄨ揪MHC鈪″瀷鎶楀師(HLA-DR)錛屾棤姝ゆ姉鍘熸彁鍛堝姛鑳藉彲浠ョ牬鍧忓厤鐤洃瑙嗕綔鐢ㄣ€?銆€銆€ 銆€銆€浜屻€佷綋娑插厤鐤?銆€銆€鐩墠琛€娓呭媯€嫻嬬粨鏋滄樉紺猴細鈶犳姉鏅氭湡錏嬬櫧鎶椾綋浜х敓鐜囦負25%-65%錛岃緝鎶楁棭鏈熻泲鐧芥姉浣撲駭鐢熺巼楂橈紱鈶℃嫻嬪嚭鐨凥PV鎶椾綋浼兼湁鍨嬬壒寮傛€э紝鏃犱氦鍙夊弽搴旓紱鈶㈡姉HPV-16E7鎶椾綋涓庡棰堢檶鐨勫瓨鍦ㄦ湁瀵嗗垏鍏崇郴錛涒懀鎶桯PV-16E4鎶椾綋涔熸槸鍙戠敓瀹鐧屻€佸鍙戞垨榪戞湡HPV鎰熸煋鐨勬爣蹇楋紱鈶や及璁℃垚浜哄拰鍎跨浜х敓IgG鎶椾綋鐨勯槼鎬х巼鐩稿悓錛屼笉鍚屽瀷鍒槼鎬х巼鍦?0%-75%涔嬮棿錛涒懃宸茶瘉鏄嶩PV-16鎴?8鍨嬭偪鐦ゆ姉浣撻槼鎬ф偅鑰呬腑錛屼粎50%-70%鍙鍑鴻鎶椾綋銆?銆€銆€ 銆€銆€涓夈€佸皷閿愭箍鐤ｈ嚜鐒舵秷閫€ 銆€銆€瀵笴A鑷劧娑堥€€灝氭棤緋葷粺璿勪及錛岀劧鑰屼互瀹夋叞鍓傚鐓х爺絀跺彂鐜板叾鑷劧娑堥€€鐜囧湪0%錛?7%銆?8%鍜?9%涔嬮棿錛孋A娑堥€€鎴栨不鎰堝悗錛屼粛鏈?5%鎮ｈ€呭瓨鍦ㄦ綔浼忔劅鏌擄紝鍏朵腑67%鎮ｈ€呭鍙戙€?銆€銆€ 銆€銆€涓村簥琛ㄧ幇 銆€銆€娼滀紡鏈?鍛ㄥ埌8涓湀錛屽鉤鍧?涓湀錛屽瑙佷簬鎬ф椿璺冪殑闈掋€佷腑騫寸敺濂籌紝鍙戠棶楂樺嘲騫撮緞涓?0-25宀侊紝鐥呯▼騫沖潎鍦?-5涓湀鐨勭敺濂蟲偅鑰咃紝鍦ㄦ€ф帴瑙﹀悗涓嶄箙鍗沖彂鐥咃紝鑰岀棶紼嬪鉤鍧?2涓湀鐨勭敺鎬ф偅鑰咃紝鍏舵€ф帴瑙﹁€呭彲涓嶅彂鐥呫€傚鏁版偅鑰呬竴鑸棤鐥囩姸銆傛崯瀹沖ぇ灝忓強褰㈢姸涓嶇瓑銆傚彲浠呬負鏁頒釜錛屼害鍙負澶氭暟閽堝ご鏍峰ぇ鐨勬崯瀹籌細鍦ㄩ槾鑲溳儴鍙暱鎴愬ぇ鐨勮偪鐦ゆ牱鐗╋紝鏈夊帇榪劅錛涙湁鎭惰嚟鍛籌紱鏈夋椂灝忕殑婀跨枺鍙嚭鐜伴槾閮ㄧ棝鐥掍笉閫傦紝鐥呬漢鍙嚭鐜板翱琛€鍜屾帓灝垮洶闅撅紱鐩磋偁鍐呭皷閿愭箍鐤ｅ彲鍙戠敓鐤肩棝銆佷究琛€錛岃€岀洿鑲犲唴澶х殑婀跨枺鍒欏彲寮曡搗閲屾€ュ悗閲嶆劅銆?銆€銆€鐢鋒€ф偅鑰呭ソ鍙戜簬鍖呯毊緋誨甫銆佸啝鐘舵矡銆佸寘鐨€佸翱閬撱€侀槾鑼庛€佽倹闂ㄥ懆鍥村拰闃村泭銆傜棶鍒濅負娣＄孩鎴栨薄綰㈣壊綺熺姸澶у皬璧樼敓鐗╋紝鎬ц川鏌旇蔣錛岄《絝◢灝栵紝閫愭笎闀垮ぇ鎴栧澶氥€傚彲鍙戝睍鎴愪鉤澶寸姸鎴栧泭鐘訛紝鍩哄簳紼嶅鎴栨湁甯︼紝琛ㄩ潰鏈夐綺掋€傚湪鑲溳棬閮ㄥ父澧炲ぇ錛岀姸濡傝彍鑺憋紝琛ㄩ潰婀挎鼎鎴栨湁鍑鴻錛屽湪棰楃矑闂村父縐瓨鏈夎創娑詫紝鏁ｅ彂鎭惰嚟姘斿懗錛屾悢鎶撳悗鍙戶鍙戞劅鏌撱€備綅浜庢箍搴﹁緝浣庡共鐕ラ儴浣嶇殑鐢熸畺鍣ㄧ枺錛屾崯瀹沖父灝忚€屽憟鎵佸鉤鐤ｇ姸銆備綅浜庢箍鐑箍娑﹂儴浣嶇殑鐤ｅ父琛ㄧ幇涓轟笣鐘舵垨涔沖ご鐦ょ姸錛屾槗鋙嶅悎鎴愬ぇ鐨勫洟鍧椼€傛湁涓ラ噸鑲濈棶鐨勬偅鑰呮箍鐤ｅ彲澧炲ぇ銆傚濞犲彲浣挎箍鐤ｅ鍙戞垨鐢熼暱鍔犲揩銆?銆€銆€浜氫複搴婃劅鏌撴槸鎸囦複搴婁笂鑲夌溂涓嶈兘杈ㄨ鐨勭棶鍙橈紝浣嗙敤3%-5%閱嬮吀娑插矓閮ㄥ娑傛垨婀挎暦5-10鍒嗛挓鍙湪HPV鎰熸煋鍖哄煙鍙戠櫧錛屽嵆鎵€璋撯€滈唻閰哥櫧鐜拌薄鈥濄€?銆€銆€HPV鎰熸煋鍜岃偪鐦ょ殑鍙戠敓錛?銆€銆€錛堜竴錛塇PV涓庣毊鑲よ偪鐦ょ殑鍙戠敓鏈夊叧 銆€銆€瀹冨湪鐨偆鐧屽拰鍏朵粬瑙ｅ墫閮ㄤ綅鐨勮偪鐦や技涔庤搗鍐沖畾浣滅敤銆傚彛鑵旇壇鎬ц禈鐢熺墿鍜岀檶鍓嶇棶鍙橈紝鐨偆槌炵姸緇嗚優鐧岀粍緇囦腑鍙彂鐜癏PV-11銆?6銆?8鍨婦NA錛屾浘鎶ラ亾鍠夐儴HPV-6涔沖ご鐦ゆ伓鍙樻垚鍠夌檶錛岀毊鑲ょ枺鐘惰〃鐨彂鑲蹭笉鑹紙EV錛夋槸HPV娼滃湪鑷寸檶浣滅敤鐨勮瘉鎹紝EV鐨崯涓彂鐜板縐岺PV鍨婦NA錛屽苟鍦ㄦ偅鑰呯毊鑲ら碁鐘剁粏鑳炵檶涓鍑篐PV-5銆?銆?4銆?7鍙?0鍨嬶紝鐨偆槌炵姸緇嗚優鐧屼技涔庢槸鐢卞厛宸插瓨鍦ㄧ殑鐥呮瘨鎬ф崯瀹蟲伓鍙樿€屾潵銆?銆€銆€錛堜簩錛夊皷閿愭箍鐤ｄ笌鑲溳棬鐢熸畺鍣ㄧ檶 銆€銆€鐢熸畺鍣ㄧ檶涓嶩PV綾誨瀷鏈変竴瀹氱殑鍏崇郴銆傚埄鐢―NA鏉備氦鎶€鏈彂鐜扮敓孌栧櫒鐧岀粍緇囦腑瀛樺湪HPV-6銆?1銆?6銆?8鍨嬬瓑銆?銆€銆€1錛庡棰堢檶錛氭牴鎹瓾PV涓庡棰堢檶鐨勫叧緋夥紝鍙皢鍏跺垎涓轟袱澶х被鍨嬶細浣庡嵄鍨嬩富瑕佹寚HPV-6銆?1鍨嬶紝楂樺嵄鍨嬫槸鎸嘓PV-16銆?8鍨嬶紝Gissmann絳夎瀵熷埌鍦ㄤ鏡琚€у棰堢檶涓紝鏈?7.4%鎮ｈ€呭瓨鍦ㄧ潃HPV-16銆?8鍨嬶紝鍏朵粬瀛﹁€呬篃鏈夌溮鍚屽彂鐜般€傝繕鏈変漢浠庝鏡琚€у棰堢檶涓垎紱誨嚭HPV-33鍜?5鍨嬨€?銆€銆€2錛庣毊鑲ら碁鐘剁粏鑳炵檶錛圫CC錛夛細HPV鎰熸煋鑰屽彂鐢熺殑CA涔熷彲鑳芥槸鐧屽墠鎹熷錛屽苟鍙彂灞曟垚鑲溳棬鐢熸畺鍣⊿CC錛岃繖琛ㄦ槑HPV鏄コ闃淬€侀槾鑼庡強鑲溳棬鐢熸畺鍣⊿CC鐨勯噸瑕佸洜緔犮€侰A錛屽法澶A鍜岀枺鐘禨CC緇勬垚涓€涓敓孌栧櫒鐧屽墠鐥呭彉鍜岀檶鐨勬崯瀹崇棶璋憋紝鏈変簺閮ㄤ綅鐢熸畺鍣ㄧ檶鐥呬緥鍦ㄥ叾鍛ㄥ洿鐨偆鏈塁A瀛樺湪錛屾湁鏃惰倝鐪艱涓哄吀鍨嬬殑CA錛屼絾緇勭粐瀛︽鏌ヤ腑鍙戠幇SCC鐨勫絝嬬棶鐏躲€?銆€銆€3錛庨矋娓╂牱涓樼柟鐥咃細甯歌浜庨槾鑼庛€佸コ闃存垨鑲溳棬鍛ㄥ洿錛屾浘鍦ㄧ毊鎹熷唴鍙戠幇HPV-16鍨婦NA銆?銆€銆€ 銆€銆€